MX2008004723A - Non-invasive in vitro method to detect transitional cell carcinoma of the bladder - Google Patents

Abstract

The present invention refers to a non-invasive in vitro method to detect a transitional cell carcinoma of the bladder in an individual via urine analysis, to determine the stage or severity of this cancer in an individual or to monitor the effect of treatment administered to an individual suffering from this cancer.

Description

IN VITRO NON INVASIVE METHOD TO DETECT CELL CARCINOMA OF THE BLADDER TRANSITION FIELD OF THE INVENTION The present invention relates to a non-invasive in vitro method for detecting the presence of transitional cell carcinoma of the bladder in the urine of an individual via urinalysis, as well as the use of peptide sequences derived from the selected proteins and a team to perform the method.

BACKGROUND OF THE INVENTION Bladder cancer is the most common cancer of the urinary tract; It is also the fourth most common cancer in men and the eighth most common in women. It includes a broad spectrum of tumors of various histological types including transitional cell carcinoma of the bladder (BTCC, 90CF), squamous cell carcinoma (7%), adenocarcinoma (2 °) and undifferentiated carcinoma (1 %) The grade and stage of the tumor are the best indicators of BTCC prognosis. Bladder tumors are cytomorphologically graduated from Gl (low grade) to G3 (high grade) in a decreasing state of differentiation and increasing aggressiveness of the disease according to the World Organization of Greeting (WHO). With respect to the stage or mvasivity, BTCCss are classified as superficial papillary (Ta and TI), invasive muscle (T2 to T4) or carcinoma m situ or tumor mite not common (TIS, for its acronym in English). Low-grade tumors are usually confined to the mucosa or infiltrated superficial layers (stage Ta and TI). Most high-grade tumors are detected at least in the TI cap (invasive lamina propria). Approximately 75% of diagnosed bladder cases are superficial. The remaining 25% are invasive of the muscle at the time of diagnosis. Patients with superficial CBCT have a good prognosis but have a 70% recurrence risk. Despite this high risk, Ta tumors tend to be of garlic grade and only 10-15% will progress to muscle invasion in 2 years. However, the percentage of IT tumors that progress to the T2 stage is higher (30-50%). Currently, the best diagnostic system for bladder cancer in individuals is established by cystoscopy and transurethral biopsy or by resection, both representing invasive procedures. Flexible cystoscopes make the technique less aggressive, but remain invasive and highly unfavorable, requiring even some form of anesthesia.

The prevalent noninvasive technique for the diagnosis of BTCC is to identify neoplastic cells by morphological examination of the cells in urine. Therefore, cytology is currently used to monitor patients diagnosed with and treated for bladder cancer. Although urine cytology can detect in situ tumors that can not be detected by cytoscopy as well as tumors located in the upper end of the bladder or upper urinary tract, ie, urethra, pelvis and kidney, several studies have shown that cytology It has a very low sensitivity in the diagnosis of bladder cancer, missing up to 50% of the tumors. The findings in cytologies are weak and can be confused with reactive degenerative processes. Cytology studies require individual expert evaluation that delays the availability of results and also introduces subjectivity and variance to the final results. Actually, there is no highly sensitive and non-invasive specific method available to diagnose bladder cancer (Boman H. and others J Urol 2002, 167: 80-83). Many efforts have been made to improve the detection of non-invasive cancer. Recent advances in the creation of the profile of the expression of cancer cells by proteomic technologies, high resolution two-dimensional electrophoresis and mass spectrometry have made it possible to identify candidate proteins as tumor markers for bladder cancer, such as nuclear matrix protein NMP22 ( Solowa YS and golds, J Uiol 1996, 156: 363-367), hyaluronic acid and hyaluronidase (Pham HT, et al., Cancer Res 1997,57: 778-783), base membrane complexes (BTA), Pode D. et al., J Urol 1999, 161: 443-446), carcinoembryonic antigen (CEA, Halim AB et al., Int J Biol Markers, 1992; 7: 234-239), uroplakin II (Wu XT et al., Cancer Res 1998; 58: 1291-1297), scattering factor / hepatocyte growth factor (SF / HGF, Gohji K. and others, J Clin Oncol 200; 18: 2963-2971), the proteins of the keratin / cytokeratin family as cytokeratin 20 (Buchumensky V. et al., J Urol 1998, 160: 1971-1974), cytokeratin 18 (Sánchez-Carbayo M et al., Clin. Cancer Res 200, 6: 2585-3594), 8-Ka protein of mammary tumor (MAT-8, Morpson BW, et al., J. Biol Chem 1995, 270: 2176-2182) and telomerase (Lee DH et al., Clinical Cancer Research 1998, 4: 535-238). Memon A. A. et al. (Cancer Detect Prev 2005, 29: 249-255) identifies seven differentially expressed proteins in bladder cancer from cell lines and biopsies. However, no control is used for comparison. Langbein, S. et al. Technol Cancer Res Tetra 2006, 67-71) describes the profile proteins of biopsies of bladder cancer patients using 2D-PAGE and SELI-TOF-MS techniques.

Rasmussen H. H. et al. (J Urol 1996, 155: 2113-2119) describes a database of most of the abundant proteins present in the urine of patients with bladder cancer. Celis J. E. et al (Electrophoresis 1999, 300-309) describes a database of proteins present in the biopsies of patients with bladder cancer. However, no markers have been identified in any of these references, since the authors do not show a quantified differential profile of proteins in healthy samples against tumor samples. Furthermore, no marker to predict the prognosis and degree of bladder cancer has been shown to have been useful in clinical trials (Miyake H. et al., J Urol 2002, 167: 1282-1287).

DESCRIPTION OF THE INVENTION The present invention provides an efficient and fast, highly sensitive in vitro non-invasive method that refers to proteins associated with BTCC. It also refers to a method for the diagnosis, prognosis and monitoring of BTCC via the analysis of urine samples. The present invention surprisingly provides biomarkers of bladder cancer for the detection of BTCC via urinalysis. A preferred embodiment of the invention relates to 40 biomarkers of bladder cancer for the detection of BTCC via urinalysis which have a significant value for the detection, diagnosis, prognosis and / or monitoring of BTCC. Am? Noac? Lasa-1 (ACX1) is a zinc-binding, homodimepca, cytosolic enzyme that catalyzes the hydrolysis of L-amino acids acylated to L-amino acids and acyl group and has been postulated to function in the catabolism and recovery of acylated amino acids . ACY1 has been assigned to chromosome 3p21.1, a region reduced to homozygosity in small cell lung cancer (SCLC), and it has been reported that its expression has been reduced or undetectable in the cell lines of SCLC and tumors. ACY1 is the first member of a new family of zinc binding enzymes. The aldehyde reductase (AKR1A1) belongs to the family aldo / keto reductase, is involved in the reduction of biogenic aldehydes and xenobiotics and is present in virtually every tissue. Aldolase (ALDOB), also called 1-phosphate aldolsa, was produced in the liver, kidneys and brain. It is needed for the decomposition of fructose. Carcinoid alpha-amylase (AMi), pancreatic alpha-amylase, and FA-amylase belong to the glycosyl hydrolase family 13 and hydrolyze 1,4-alpha-glucoside bonds in oligosaccharides and polysaccharides and therefore catalyzes the first step in digestion of dietary starch and glycogen. The human genome has a set of several amylase genes that are expressed at high levels in the salivary gland or pancreas. Anexma IV (ANXA4) belongs to the anexma family of calcium-dependent phospholipid binding proteins. Although its ammonium functions have been clearly defined, several members of the annexa family have been implicated in events related to membranes along with exocytotic and endocytotic routes. ANXA4 are almost exclusively expressed in epithelial cells. Apolipoprotein Al (APOA1) belongs to the apolipoprotein A1 / A4 / E family and participates in the reverse transport of cholesterol from tissues to the liver for secretion by promoting the flow of cholesterol from tissues and acting as a cofactor for cholesterol acyltransferase. lecithin (LCAT, for its acronym in English). Biliverdin reductase A (BIEA) belongs to the gfo / idh / moca family and the biliverdma reductase subfamily and reduces the gamma-ethene bridge of open tetrapyrol, biliverdin IX alpha, to bilirubin with the concomitant oxidation of NADH or NADPH. Flavin reductase (BLVRB) catalyses the transfer of electrons from reduced indin nucleotides to flavins as well as to methylene blue, quark pyrroliquinoline, riboflavin or metemoglobin. BLVRB has a possible role to protect cells from oxidite damage or to regulate iron metabolism. In the liver, converts biliverdin to bilirubin. It is expressed predominantly in the liver and erythrocytes, and at levels in the heart, lung, adrenal gland and brain. Cathepsin D (CTD) is an acid protease that belongs to the Al peptidase family and activates the decomposition of the intracellular protein. It is involved in the pathogenesis of several diseases such as bladder cancer (Ozer E., et al., Urology 1999, 54: 50-5 and E. loaquima, and others, Anticancer Res 2002, 22: 3383-8), cancer. breast and possibly Alzheimer's disease. Cathepsin D is synthesized as an inactive 52 kDa precursor, consisting of two 14 kDa polypeptides (CATD H) and one 34 kDa polypeptide (cATD K). The inactive peptide is activated by proteolysis of the N-terminus resulting in the 48 kDa enzymatically active protein. The complement precursor C3 (C03) plays a central role in the activation of the complement system. Its processing by C3 convertase is the central reaction in both complement pathways, C03 has been linked to urogenital tumors (Dunzendorfer U., et al., Eur Urol 1980, 6: 3232-6).

Dipeptidase does not specify cytosolic (CPGL1) belongs to the family of peptidase M20A and is also known as tissue of camosmase and peptidase A. It is a non-specific dipeptidase instead of a selective camosmase. Alpha Enolase (ENOA) is a multifunctional enzyme that, like its role in glycolysis, plays a part in several processes such as growth control, tolerance to hypoxia and allergic responses. It can also function in the pericellular intravascular and fibrinolytic system due to its ability to serve as a receptor and activator of plasminogen on the cell surface of various cell types such as leukocytes and neurons. The enolase of mammals is composed of 3 subunits of isozyme, alpha, beta and gamma, which can form homodimers or heterodimers that are of the cell type and specific for development. ENOA interacts with PLG in the neuronal plasma membrane and promotes its activation. It is used as a diagnostic marker for many tumors and, in the heterodimeric form, alpha / gamma, as a marker of hypoxic brain damage after cardiac arrest. It has been linked to bladder cancer (Iczko ski K.A., et al., Histoplogy 1999, 35: 150-6). The ferpnite light chain (FRIL) belongs to the ferritma family and is the main intracellular iron storage protein in prokaryotes and eukaryotes.

It is composed of 24 subunits of heavy and light ferritin chains. Variation in the composition of the ferritm subunit may affect iron absorption and release rates in different tissues. A main function of ferptin is the storage of iron in a soluble and non-toxic state. The Rab GDP beta dissociation inhibitor (GDIB) belongs to the gdi rab family and regulates the GDP / GTP exchange reaction of family members, small GTP binding proteins of the superfamily which is involved in the vesicular traffic of molecules between cellular organelles. GDIB is ubiquitously expressed. Plasma glutone peroxidase (GPX3) belongs to the glutone peroxidase family and catalyzes the reduction of hydrogen peroxide, organic hydroperoxide, and lipid peroxides by reduced glutone and works in the protection of cells against oxidative damage. It has been shown that human plasma glutone peroxidase is an enzyme containing setenium. The expression of GX3 seems to be tissue-specific. The glutone synthetase (GSHB) is important for a variety of biological functions, including cell protection from oxidative damage by free radicals, detoxification of xenobiotics and membrane transport.

Gelsolin (GSN40 and GSN80) is an actma modulating protein, regulated with calcium that binds to the additional ends of actin monomers or filaments, preventing the exchange of monomers (blocking or end capping). It can promote the assembly of monomer sen filaments (nucleation), as well as existing filaments of the server. In addition, this protein binds to actin and fibrilnectom. It has been related to bladder cancer, although it has not been described as a bio-burn, that is, its level of expression has not been compared and quantified in healthy individuals against cancer patients (Rasmussen, HH et al., J Urol 1996, 155 : 2113-2119; Do K. Hokkaido Igaku Zasshi 2003, 78: 29-37; Celis A et al., Electrophoresis 1999, 20: 355-61). Glutone S-transferase (GSTP1) belongs to a family of enzymes that play an important role in detoxification by catalyzing the conjugation of many hydrophobic and electrophilic compounds with reduced glutone. The dehydrogenated cytoplasmic isocitrate (IHC) belongs to the family of isocitrate and isopropyl malate dehydrogenase and catalyzes the oxidative decarboxylation of isocitrate to form alpha-ketoglutarate. The vesicular integral membrane protein precursor (VIPP36 (LMNN2) plays a role as an intracellular lectin in the early secretory pay, interacts with N-acetyl-D-galatosamine and glycans of the high-mannine type and can also bind to 0-lyzed glycans It is involved in the transport and classification of glycoproteins that carry glycans of the high-mannose type.6 The antigen complex 6 of lymphocytes, site 6 G6E (LY6G6E) belongs to the amphiano-6 superfamily of Lmfocitos. The members of this family are rich in cistern, generally cellular surface proteins anchored with GPI, which have definite or putative immune-related roles. Its specific function is unknown. The morning-binding lectin sepna protease 2 precursor (MaSP2) belongs to the peptidase family if and is a trypsin protease that presumably plays an important role in the initiation of the mannose binding lectin complement activation pathway ( MBL). After activation, it separates C4 that generates C4A and C4B. Nicotinate nucleotide pyrophosphotylase (NADC) belongs to the nadC / mod D family and is an intermediate in the de novo NAD synthesis pathway of tryptophan and acts as a potent endogenous excitotoxin through the hyperstimulation of N-methyl receptor D-aspartate (NMDA, for its acronym in English) in the neuron. Elevation of NADC levels in the brain has been linked to the pathogenesis of neurodegenerative disorders such as epilepsy, Alzheimer's disease and Huntington's disease. It has not yet been linked to cancer. Napsina A (NAPSA) belongs to the peptidase family al and is expressed at the highest levels in the kidney, at a moderate level Jan. lung and at low levels in the spleen and adipose tissue. It may be involved in the process of pneumocyte surfactant precursors. The protein associated with 2G4 proliferation (PA2G4) is a single double-stranded DNA binding protein that shows the specific variation of the cell cycle in nuclear localization, which belongs to the m24c family of peptidase. As the protein is expressed in response to mitogen stimulation, it may belong to a large family of cell cycle regulatory proteins or replication proteins that maintain the cell cycle activities of proliferating cells. The DJ1 oncogene (PaRK7) is highly expressed in pancreas, kidney, skeletal muscle, liver, testes and heart and acts as a positive regulator of androgen receptor dependent transcription. It can function as a redox sensitive chaperon and as a sensor for oxidative stress. It prevents the aggregation of SNCA and protects neurons against oxidative stress and cell death.

The pol? (Rc) 1 binding protein (PCRPB1) is a nucleic acid binding protein of a strand that binds preferably to oligo dC. It can shoot between the nucleus and the cytoplasm. It is abundantly expressed in the skeletal muscle, the thymus and peripheral blood leukocytes while an inferior expression is observed in the prostate, spleen, testes, ovary, small intestine, heart, liver, adrenal and thyroid glands. The programmed cell death interaction protein 6 (PDC6I) is expressed in the brain, heart, placenta, lung, kidney, pancreas, liver, skeletal muscle or shock and may play a role in the regulation of apoptosis and cell proliferation. Interact with PDCD6 / ALG-2 and SH3KBP1. The lysosomal acid phosphatase precursor (PPAL) belongs to the family of histidine acid phosphatase and hydrolyzes orthophosphoric monoesters to alcohol and phosphate. Peroxiredoxin 2 (PRDX2) belongs to the ahpc / tsa family and is involved in the regulation of redox of the cell. Reduce peroxides with the reduction equivalents provided through the thioredoxin system. It can not receive electrons from glutaredoxin. It can play an important role in the elimination of peroxides generated during metabolism. It can participate in the growth factor signaling cascade and tumor necrosis factor alpha by regulating the intracellular concentrations of H202. Increases the activity of natural killer cells (NK). Glutamyl peptide cyclotransferase (QPCT) is responsible for biosynthesis of pyroglutamyl peptides. It has an inclination against acid residues and tryptophan adjacent to an N-terminal glutamic acid. The plasma retinol binding protein (RETBP) belongs to the lipocalcmal family and is the specific carrier for retinal (vitamin A alcohol) in the blood. Supplies the retour of the liver stores to the peripheral tissues. In plasma, the RBP-retinol complex interacts with transtiretma, which prevents its loss by filtration through the kidney glomeruli. A vitamin A deficiency blocks the secretion of the binding protein post-translationally and results in defective delivery and provides the epidermal cells. The selenium binding protein (SBP1) belongs to the selenium binding protein family. Selenium is an essential nutrient that exhibits potent anticarcinogenic properties; A deficiency in setenium can cause certain neurological diseases. It has been proposed that the effects on selenium to prevent cancer and neurological diseases can be mediated by selenium binding proteins. Stress-induced phosphoprotein 1 (STIP1) is an adapter protein that mediates the association of molecular chaperones HSC70 and HSP90 (HSPCA and HSPCB). Transaldolase (TALDO) belongs to the family of transaldolase and is a key enzyme in the non-oxidative pentose phosphate pathway that provides r-biose-5-phosphate for nucleic acid synthesis and NADPH for lipid biosynthesis. This route can also maintain glutathione in a reduced state and therefore protects the sulfohydryl groups and cellular integrity of oxygen radicals. Transtiretma (TTHI, micially prealbumin) is one of 3 thyroid hormone binding proteins found in the blood of vertebrates. It is produced in the liver and circulates in the bloodstream, where it binds to retinol and thyroxine. The WD 1 repeat protein (WDR1) belongs to the repeating wd aipl family and induces the disassembly of actin filaments together with the ADF / cofilma family proteins. Cancer can be detected by analyzing the expression pattern of at least one biomarker of bladder cancer in a test urine sample. Therefore, the detection of at least one protein differentially expressed dwarf urine test sample compared to normal urine indicates BTCC. Urine samples are very diverse in their composition. Thus, normalization is essential to take into account differences in total protein concentration and to remove the tilt from sample to sample. The expression levels of a control protein whose urine content is always constant can be used to normalize signal levels. In the literature there are many examples of these non-variable proteins. In the present invention, it was shown that the transferin is a non-variable protein. Representative bladder cancer proteins or biomarkers identified in the present invention include, but are not limited to, TTHY, ACY1, AKR1A1, ALDOB, ANXA4, BIEA, BLVRB, CATD H, CATD K, C03, CPGL1, ENOA, FRIL, GDIB, GPX3, GSHB, GSTP1, IDHC, LMAN2, LY6G6E, MASP2, NADC, NAPSA, PA2G4, PARK7, PCBP1, PDC6I, PPAL, PRDX2, PTD012, QPCT, SBP1, STIP1, TALDO, WDR1, AMY, APOA1, GSN40, GSN80 and RETBP (also referred to as biomarkers of bladder cancer 40). One aspect of the present invention relates to a non-invasive in vitro method comprising: a) detecting and quantifying one or more biomarkers present in an urine test sample of an individual; and b) comparing the measurement of expression obtained in a) in the test urine sample to the corresponding standard value in normal urine, where variations in the measurement obtained in a) compared to the corresponding standard value in normal urine indicate BTCC. This method can be adapted for screening to detect the presence of BTCC, in order to determine the stage or severity of this cancer. In addition, this method can also be used to assess the lack of disease after surgical resection, to establish the diagnosis and / or prognosis of this cancer and / or to monitor the effect of the treatment administered to an individual suffering from said cancer. Another aspect of the invention relates to the use of one or more peptide sequences derived from biomarkers present in urine to detect the presence of BTCC via urinalysis, to determine the stage or severity of this cancer, to assess the lack of disease after of surgical resection, to establish the diagnosis and / or prognosis of this cancer and / or to monitor the effect of the treatment administered to an individual suffering from said cancer. Another aspect of the invention relates to the use of one or more nucleotides or peptide sequences derived from the biomarkers of bladder cancer or a transcriptional or post-translational variant, alone or in any combination, in methods for screening, identifying, developing and evaluate the efficacy of therapeutic agents to treat BTCC.

The invention further provides antibodies directed against said biomarkers of bladder cancer. These antibodies may be provided in a variety of forms, as appropriate, for a particular use, including, for example, in a soluble form, immobilized on a substrate or in combination with a pharmaceutically acceptable carrier. Another aspect of the invention relates to a kit for performing the method as defined above comprising 1) at least one antibody that specifically recognizes a cancer biomarker in urine and 2) a vehicle in a suitable package. Another aspect of the invention relates to a reference expression profile of BTCC comprising an expression pattern of one or more biomarkers of bladder cancer. Different reference expression profiles of BTCC can be established and correlated to different stages of cancer. For the purposes of the present invention, the following definitions have been used: The term "cancer" refers to the disease that is typically characterized by growth of abnormal or unregulated cells capable of invading adjacent tissues and spreading to distant organs. The term "carcinoma" refers to tissue that results from the growth of abnormal and unregulated cells. The term "transitional cell carcinoma of the bladder" or if abbreviation "BTCC" refers to any malignant proliferative disorder in bladder epithelial cells. The term "tumor" refers to any abnormal mass of tissue generated by a neoplastic process, where that is benign (non-cancerous) or malignant (cancerous). The term "bladder cancer proteins or biomarkers" refers to proteins differentially expressed in BTC, ie, proteins that are differentially expressed in the urine of a healthy individual compared to the urine of a patient having BTCC. In the present invention the group of proteins comprised of differentially expressed proteins includes the biomarkers selected from the group comprising TTHY, ACY1, AKR1A1, ALDOB, ANXA4, BIEA, BLVRB, CATD H, CATD K, C03, CPGL1, ENOA, FRIL, GDIB, GPX3, GSHB, GSTP1, IDHC, LMAN2, LY6G6E, MASP2, NADC, NAPSA, PA2G4, PARK7, PCBP1, PDC6I, PPAL, PRDX2, PTD012, QPCT, SBP1, STIP1, TALDO, WDR1, AMY, APOAl, GSN40, GSN80 and RETBP, as well as any protein in a urine sample whose ratio changes at least twice when compared to two different urine samples: one from a healthy individual and not from a transitional cell carcinoma patient, is quantification being carried out by an image analyzer software, for example Progenesis PG220 software. Biomarkers of bladder cancer as described herein may be of any length and further comprise additional sequences derived from the native protein and / or heterologous sequences, any trans-translational or post-translational variant, as well as any sequence with at least one 95% identity with the sequences described, where identity is defined as the percentage of residues that are identical between two sequences. Those skilled in the art will appreciate that these other portions or variants may also be useful in the treatment and detection of cancer. The term "detecting one or more biomarkers" means determining the existence of one or more biomarkers, while the term "quantification of one or more biomarkers" means expressing the presence of one or more biomarkers as a value / for example as a value of intensity). The term "indicative of BTCC" means the variations found in the measurement of one or more biomarkers in a urine sample test compared to the corresponding normal value in normal urine serves as a test for the presence of BTCC. The term "variation" refers to a change in the level of expression of a protein. The term "differentially expressed protein" refers to a protein, whose expression pattern varies (increases or decreases) in the urine of a patient who has BTCC purchased with the urine of a healthy individual. The term "inversely expressed" refers to two differentially expressed proteins whose expression patterns are or are opposite (i.e., in one sample one is overexpressed while the other is repressed or vice versa). The term "protein extract" corresponds to the supernatant obtained after centrifugation of the urine sample. The term "individual" refers to all animal species classified as mammals and includes, but is not restricted to, domestic and farm animals, primates and humans and preferably refers to a woman or man of any age or race. The term "healthy individual" refers to an individual who does not suffer from BTCC and may include patients who have other urological diseases. The term "previously diagnosed" refers to an individual who has already received a first positive diagnosis for BTCC. The term "previously undiagnosed" refers to an individual who has never received a positive first diagnosis for BTCC (de novo diagnosis.) The term "standard value in normal urine" refers to the quantification of the average expression level of one or more biomarkers detected in a single urine sample of individuals known to not suffer from BTCC.

The term "diagnosis" of BTCC refers to the process to identify or determine the nature and cause of BTCC through the evaluation of one or more BTCC biomarkers. The term "prognosis" refers to the probable outcome or course of a disease that is the opportunity for recovery or recurrence. The term "monitor" means to assess the presence or absence of BTCC in an indifferent individual moment. The term "treatment" refers to any process, action, application or the like, wherein that subject an individual to medical help in order to improve their condition, directly or indirectly. The term "specificity" refers to the ability of a test to exclude the presence of a disease when it is not actually present. Specificity is expressed as the number of healthy individuals for whom there is a correct negative test (this group being called real negatives), divided by the sum of the real negatives and the number of healthy individuals for whom there is an incorrect positive test (this group being called false positives). The term "sensitization" refers to the ability of a test to detect a disease when it is actually present. Sensitivity is expressed as the number of sick patients for whom there is a correct positive test of that group being called false negatives). The term "robustness" defines the ability of the numerical method to provide the same result despite the variability in the initial samples. The term "gene" refers to a region of a molecular chain of desoxybonucleotides that encodes a protein and may represent a portion of a coding sequence or a complete coding sequence. The term "protein" refers to at least one molecular chain of amino acid mter linked olecularly through covalent or non-covalent bonds. The term includes all forms of post-translational protein modifications, for example glycosylation, phosphorylation or acetylation. The terms "peptide" and "polypeptide" refer to molecular chains of amino acids that represent a protein fragment. The terms "protein" and "peptide" are indistinguishable. The term "antibody" refers to a Y-shaped protein (known as immunoglobulin) on the surface of B cells that is secreted in the blood or lymph in response to an antigenic stimulus, such as an exogenous protein, bacteria, virus, parasite or organ transplanted, and that exhibits a specific binding activity for a white molecule called an "antigen". The binding region of immunoglobulin antigens can be divided into F (ab ') 2 fragments of Fab. The term "antibody" includes monoclonal and polyclonal antibodies, whether intact or fragments derived therefrom; and includes human antibodies, humanized antibodies and antibodies of non-human origin. A "non-human antibody" is an antibody generated by an animal species other than Homo sapiens. A "humanized antibody" is a genetically-treated antibody in which the minimal mouse part of a murine antibody is fused to a human antibody. Generally, humanized antibodies are 5-10% mice and 90-95% human. A "human antibody" is an antibody derived from a transgenic mouse that carries genes from human antibodies or from human cells. "Polyclonal antibodies" include populations of heterogeneous antibodies that target different antigenic determinants of the target molecule. The term "specific antibody" refers to an antibody raised against a specific protein (in this case against a particular bladder cancer marker). The term "antibody-protein complex" refers to a complex formed by an antigen and its specific antibody. The term "body-cornbi" (combinatorial antibody) refers to an antibody exhibited in filamentous phages, which allows the direct screening of cDNA libraries for the expression of reactive antibodies on its cell surface., without the need for production and purification of antibodies using bacterial or eukaryotic cell systems. The term "Fab recombinant antibody" refers to a recombinant antibody that only contains the Fab fragment that is univalent and useful when the antibody has a very bad affinity for its antigen. They can be obtained recombinantly if the protein sequence is known. The term "ScFV antibody fragment" refers to a single chain variable fragment (scFv) that can be expressed in bacterial cultures. The term "epitope" refers to an antigenic determinant of a protein, which is the amino acid sequence of the protein that recognizes a specific antibody. Said epitopes can be comprised of a contiguous extension of amino acids (linear epitope) or of non-contiguous amino acids that come between each other by virtue of the three-dimensional fold of the polypeptide chain (discontinuous epitopes). The term "solid phase" refers to a non-aqueous matrix to which the antibody can bind. Examples of materials for the solid phase include but are not limited to glass, polysaccharides (for example random), polyacrylamide, polystyrene, polyvinyl alcohol and silicones. Examples of solid phase forms are the well of a plate or a purification column. The term "submersion strip" refers to a device immersed in a liquid to perform some kind of test that can determine and / or quantify some properties of the liquid (chemical, physical, etc.). This kind of submersion strip is usually made of paper or cardboard and is impregnated with reagents whose color changes indicate some aspect of the liquid. The term "vehicle" refers to a mechanism or device by which something is transported or driven. The term "packaging" refers to the content and packaging before the sale with the main purpose of facilitating the purchase and use of a product. The term "biomicrocircuit" refers to the collection of miniaturized test sites (microdispositions) arranged on a solid substrate that allows many tests to be carried out at the same time in order to achieve a higher rate and speed. In one embodiment of the invention, the first step of the method for evaluating bladder cancer comprises measuring one or more biomarkers selected from TTHY, AC? L, AKR1A1, ALDOB, ANXA4, BIEA, BLVRB, CATD H, CATD K, C03, CPGL1 , ENOA, FRIL, GDIB, GPX3, GSHB, GSTP1, IDHC, LMAN2, LY6G6E, MASP2, NADC, NAPSA, PA2G4, PARK7, PCBPl, PDC6I, PPAL, PRDX2, PTD012, QPCT, SBPl, STIP1, TALDO, WDRl, AMi , APOAl, GSN40, GSN80 and RETBP (also referred to as the 40b? Markers of bladder cancer) or a transcriptional or post-translational vanant thereof in an individual's urine test sample. In another embodiment, the first step of the method for evaluating bladder cancer comprises measuring one or more selected biomarkers of TTHY, ACY1, AKR1A1, ALDOB, ANXA4, BIEA, BLVRB, CATD H, CATD K, C03, CPGL1, ENOA, FRIL. , GDIB, GPX3, GSHB, GSTP1, IDHC, LMAN2, LY6G6E, MASP2, NADC, NAPSA, PA2G4, PARK7, PCBP1, PDC6I, PPAL, PRDX2, PTD012, QPCT, SBP1, STIP1, TALDO and WDR1 or a transcriptional variant or post -translational of the same dwarf sample of urine test of an individual. In another embodiment, the first step of the method for evaluating bladder cancer comprises measuring at least two biomarkers or a transcriptional or post-translational variant thereof in the urine sample of the test. In another embodiment, the first step of the method for evaluating bladder cancer comprises measuring at least three biomarkers or a trans-transcriptional or post-translational vanant in the test urine sample. In another embodiment, the first step of the method for evaluating bladder cancer comprises measuring at least four biomarkers or a trans-transcriptional or post-translational variant thereof in the test urine sample. In another embodiment, the first step of the method for evaluating bladder cancer comprises measuring at least five biomarkers or a transcriptional or post-translational variant thereof in the urine test. In another modality, the first step of the method for evaluating bladder cancer comprises measuring one or more selected biomarkers of TTHY, ACyl, AKR1A1, ANXA, BIEA, BLVRB, CATD H, CATD K, C03, CPGL1, ENOA, FRIL, GDIB, GPX3, GSHB , GSTP1, IDHC, LY6G6E, MASP2, NADC, NAPSA, PA2G4, PARK7, PCBP1, PRDX2, PTD012, QPCT, SBP1, STIP1, AMY, APOAl, GSN40, GSN80 and RETBP or a trans-transcriptional or post-translational variant thereof in the urine test sample. In another embodiment, the first step of the method for evaluating bladder cancer comprises measuring one or more biomarkers selected from TTHY, ACY1, AKR1A1, ANXA, BIEA, BLVRB, CATD H, CATD K, C03, CPGL1, ENOA, FRIL, GDIB, GPX3, GSHB, GSTP1, IDHC, LY6G6E, MASP2, NADC, NAPSA, PA2G4, PARK7, PCBP1, PRDX2, PTD012, QPCT, SBP1 and STIP1 or a transcriptional or post-translational variant thereof in the test urine sample. In another embodiment, the first step of the method for evaluating bladder cancer comprises measuring one or more biomarkers selected from TTHY, CATD H, CATD K, ENOA, FRIL, GSHB, GSTP1, IDCH, MASP2, PRDX2, AMY, APOAl, GSN40, GSN80 and RETBP or a transcriptional or post-translational variant in the test urine sample. In another embodiment, the first step of the method for evaluating bladder cancer comprises measuring one or more biomarkers selected from TTHi, CATD H, CATD K, ENOA, FRIL, GSHB, GSTP1, IDCH, MASP2 and PRDX2 or a transcriptional or post-transcriptional variant. translational sample in the urine sample.

In another embodiment, the first step of the method for evaluating bladder cancer comprises measuring one or more selected biomarkers of TTHY, AMY, APOAl, GSN40 and GSN80 or a transc iptional or post-translational variant in the test urine sample. In another embodiment, the first step of the method for evaluating bladder cancer comprises measuring APOAl and RETBP and GSN80 or a transcriptional or post-translational variant in the test urine sample. In another embodiment, the first step of the method for evaluating bladder cancer comprises measuring AMY, APOAl and GSN40 and GSN80 or a transcriptional or post-translational variant in the test urine sample. In another embodiment, the first step of the method for evaluating bladder cancer comprises measuring AMY, APOAl and ENOA and GSN80 or a transcriptional or post-translational variant in the test urine sample. In another embodiment, the first step of the method for evaluating bladder cancer comprises measuring APOAl, GSN40 and TTHY or a transcriptional or post-translational variant in the test urine sample. In another embodiment, the first step of the method for evaluating bladder cancer comprises measuring CATD K, GSN80 and IDHC or a transcourse or post-translational vanant in the test urine sample.

In another embodiment, the first step of the method for evaluating bladder cancer comprises measuring AMY, APOAl, GSN40 and IDHC or a transcriptional or post-translational variant in the test urine sample. In another embodiment, the first step of the method for evaluating bladder cancer comprises measuring APOAl, GSN40, GSN80 and TTHY or a transcriptional or post-translational variant in the test urine sample. In another embodiment, the first step of the method for evaluating bladder cancer comprises measuring CATD H, ENOA, GSTP1 and PRDX2 or a transcpptional or post-translational variant in the test urine sample. In another embodiment, the first step of the method for evaluating bladder cancer comprises measuring CATD K, ENOA, PRDX2 and TTHY or a transcriptional or post-translational variant in the test urine sample. In another embodiment, the first step of the method for evaluating bladder cancer comprises measuring AMY, CATD K, ENOA, IDHC and PRDX2 or a transcriptional or post-translational variant in the test urine sample. In another embodiment, the first step of the method for evaluating bladder cancer comprises measuring AMY, APOAl, GSN40, GSN80 and TTHY or a transcriptional or post-translational variant in the test urine sample.

In another embodiment, the first step of the method for evaluating bladder cancer comprises measuring CATD H, ENOA, GSTP1, MASP2, PRDX2 and TTHY or a transcriptional or post-translational variant in the test urine sample. In another embodiment, the first step of the method for evaluating bladder cancer comprises measuring AMY, APOAl, CATD K, ENOA, IDHC, PRDX2 and TTHY or a transcriptional or post-translational variant in the test urine sample. In another embodiment, the first step of the method for evaluating bladder cancer comprises measuring AMY, APOAl, CATD K, ENOA, IDHC, PRDX2 and TTHY or a transcriptional or post-translational variant in the test urine sample. In another embodiment, the first step of the method for evaluating bladder cancer comprises measuring one or more biomarkers selected from AMY, APOAl, GSN40, GSN80 or a transcriptional or post-translational variant thereof and one or more selected biomarkers of TTHY, ACY1, AKR1A1, ALDOB, ANXA4, BIEA, BLVRB, CATD H, CATD K, C03, CPGL1, ENOA, FRIL, GDIB, GPX3, GSHB, GSTP1, IDHC, LMAN2, LY6G6E, MASP2, NADC, NAPSA, PA2G4, PARK7, PCBP1, PDC6I, PPAL, PRDX2, PTD012, QPCT, SBP1, STIP1, TALDO, WDR1 and RETBP or a transcourse or post-translational vanant thereof in the test urine sample. In another embodiment, the first step of the method for evaluating bladder cancer comprises measuring one or more biomarkers selected from AMY, APOAl, GSN40, GSN80 or a transcriptional or post-translational variant thereof and one or more selected biomarkers of TTHY, ACY1, AKR1A1, ALDOB, ANXA4, BIEA, BLVRB, CATD H, CATD K, C03, CPGL1, ENOA, FRIL, GDIB, GPX3, GSHB, GSTP1, IDHC, LMAN2, LY6G6E, MASP2, NADC, NAPSA, PA2G4, PARK7, PCBP1, PDC61, PPAL , PRDX2, PTD012, QPCT, SBP1, STIP1, TALDO and WDR1 or a transcriptional or post-translational vanant in the test urine sample. In another embodiment, the first step of the method for evaluating bladder cancer comprises measuring one or more selected biomarkers of CATD H, CATD K, ENOA, GSHB, GSTP1, IDHC, PRDX2 and TTHY or a transcriptional or post-translational variant ther one or more selected biomarkers of ACY1, AKR1A1, ALDOB, ANXA4, BIEA, BLVRB, C03, CPGL1, FRIL, GDIB, GPX3, LMAN2, LY6G6E, MASP2, NADC, NAPSA, PA2G4, PARK7, PCBP1, PDC6I, PPAL, PTD012, QPCT, SBP1, STIP1, TALDO, WDR1, AMY, APOAl, GSN40, GSN80 and RETBP or a transcriptional or post-translational variant therin the test urine sample. In another embodiment, the first step of the method for evaluating bladder cancer comprises measuring one or more biomarkers selected from TTHY, CATD H, CATD K, ENOA, GSTP1, IDHC, MASP2, PRDX2, AMY, APOAl, GSN40, GSN80 and RETBP or a transcriptional or post-translational variant ther Another embodiment refers to the noninvasive in vitro method which comprises quantifying one or more relationships between two different biomarkers selected from 40 biomarkers of bladder cancer or a transcriptional or post-translational variant of the same dwarf urine sample from an individual's test and purchasing the measurement obtained in the test urine sample at the corresponding standard value in normal urine, where the variations indicate BTCC. In another embodiment, the method allows the progression of the disease to be determined when the same protein or proteins are compared from different samples obtained from the same patient at different times within the evolution of BTCC. In another embodiment, one or more of the biomarkers of the invention can be used to monitor the efficacy of pharmacological or surgical treatment. In another modality, the sample that will be analyzed is obtained from an individual who had not previously been diagnosed with BTCC. In another modality, the sample that will be analyzed is obtained from an individual who has previously been diagnosed with BTCC.

In another modality, the sample that will be analyzed is obtained from an individual who is currently receiving treatment against BTCC. In another embodiment, the method comprises obtaining an extract of proteins from the sample. The measurement usually comprises spectrometry or immunoassay. Spectrometry is usually a mass spectrometry of laser desorption / ionization with a normally increased surface or desorption mass spectrometer / assisted matrix laser imaging (MALDI). The immunoassay includes Western analysis, ELISA (enzyme-linked immunosorbent assay), RIA (radioimmunoanalysis), Competitive EIA (Immunoassay of Competitive Enzyme). DAS-ELISA (ELISA-Double Antibody Sandwich), immunocytochemical or immunohistochemical techniques, techniques based on the use of biomicrocircuits of microarray proteins including specific antibodies, analysis based on colloidal gold precipitation in formats such as submersible strips; or by chromat techniques affinity graphed, ligand binding analysis or lectin binding analysis. In another embodiment, the method for detecting cancer comprises contacting a urine sample with a molecule that specifically binds to a bladder cancer biomarker and quantifying this binding. In another embodiment, the detection and quantification of proteins comprises a first step, in which the protein extract of the sample is contacted with a composition of one or more specific antibodies against one or more epitopes of the protein or proteins, and a second step, in which the complexes formed by antibodies and proteins are quantified. In another embodiment, the specific antibodies used for the detection of proteins are of human, humanized or non-human origin and are selected from monoclonal or polyclonal antibodies, intact or recombinant fragments of antibodies, body-cornbi and Fab or antibody fragments from scFv Another embodiment refers to the use of one or more peptide sequences derived from selected biomarkers of TTHY, ACY1, AKR1A1, ALDOB, ANXA4, BIEA, BLVRB, CATD H, CATD K, C03, CPGL1, ENOA, FRIL, GDIB, GPX3 , GSHB, GSTPl, IDHC, LMAN2, LY6G6E, MASP2, NADC, NAPSA, PA2G4, PARK7, PCBP1, PDCßl, PPAL, PRDX2, PTD012, QPCT, SBPl, STIP1, TALDO, WDRl, AMY, APOAl, GSN40, GSN80 and RETBP or a transcriptional or post-translational variant thereof, wherein the biomarkers are present in urine.

Another embodiment of the invention relates to the use of one or more peptide sequences derived from the biomarkers selected from TTHY, ACY1, AKR1A1, ALDOB, ANXA4, BIEA, BLVRB, CATD H, CATD K, C03, CPGL1, ENOA, FRIL, GDIB, GPX3, GSHB, GSTP1, IDHC, LMAN2, L6G6E, MASP2, NADC, NAPSA, PA2G4, PARK7, PCBP1, PDC6I, PPAL, PRDX2, PTD012, QPCT, SBP1, STIP1, TALDO and WDR1 or a transcourse vanishing or post-translational thereof, where the biomarkers are present in urine. Another embodiment of the invention relates to the use of one or more peptide sequences derived from cancer biomarkers, wherein variations in the measurement of these dwarf biomarkers shows urine test compared to the corresponding standard value in normal urine indicates BTCC . In another embodiment, at least two biomarkers present in urine or a transcriptional or post-translational variant are used. In another embodiment, at least three biomarkers present in urine or a trans-transcriptional or post-translational variant thereof are used. In another modality at least four biomarkers present in urine or a transcppcional or post-translational variant are used. In another embodiment, at least five biomarkers present in urine or a trans-transcriptional or post-translational variant thereof are used.

Another embodiment of the invention relates to the use of one or more peptide sequences derived from biomarkers selected from TTH ?, AC? L, AKR1A1, ANXA4, BIEA, BLVRB, CATD H, CATD K, C03, CPGL1, ENOA, FRIL , GDIB, GPX3, GSHB, GSTP1, IDHC, LY6G6E, MASP2, NADC, NAPSA, PA2G4, PARK7, PCBP1, PRDX2, PTD012, QPCT, SBP1, STIP1, AMY, APOAl, GSN40, GSN80 and RETBP or a transcriptional variant or post -translational where the biomarkers are present in urine. Another embodiment of the invention relates to the use of one or more peptide sequences derived from the biomarkers selected from TTHY, ACY1, AKR1A1, ANXA4, BIEA, BLVRB, CATD H, CATD K, C03, CPGL1, ENOA, FRIL, GDIB, GPX3, GSHB, GSTP1, IDHC, LY6G6E, MASP2, NADC, NAPSA, PA2G4, PARK7, PCBP1, PRDX2, PTD012, QPCT, SBP1 and STIP1 or a transcriptional or post-translational variant thereof, wherein the biomarkers are present in urine . Another embodiment of the invention relates to the use of one or more peptide sequences derived from the biomarkers selected from TTHY, CATD H, CATD K, ENOA, FRIL, GSHB, GSTP1, IDCH, MASP2, PRDX2, AMY, APOAl, GSN40, GSN80 and RETBP, or a transcppcional or post-translational variant thereof, wherein the biomarkers are present in urine.

Another embodiment of the invention relates to the use of one or more peptide sequences derived from biomarkers selected from TTHY, CATD H, CATD K, ENOA, FRIL, GSHB, GSTP1, IDCH, MASP2 and PRDX2 or a transcourse or post-translational vanant from same, where the biomarkers are present in urine. Another embodiment of the invention relates to the use of one or more peptide sequences derived from selected biomarkers of TTHY, AMY, APOAl, GSN40 and GSN80 or a transcppcional or post-translational variant thereof, wherein the biomarkers are present in urine. Another embodiment of the invention relates to the use of peptide sequences derived from APOAl and RETBP or a trans-transcriptional or post-translational variant thereof, wherein the biomarkers are present in urine. Another embodiment of the invention relates to the use of peptide sequences derived from AMY, APOAl and GSN40 or a trans-transcriptional or post-translational variant thereof, wherein the biomarkers are present in urine. Another embodiment of the invention relates to the use of peptide sequences derived from AMY, APOAl and ENOA or a trans-transcriptional or post-translational variant thereof, wherein the biomarkers are present in urine. Another embodiment of the invention relates to the use of peptide sequences derived from APOAl, GSN40 and TTHY or a transcriptional or post-translational variant thereof, wherein the biomarkers are present in urine. Another embodiment of the invention relates to the use of sequences ^ of peptides derived from CATD K, GSN80 and IDHC or a transcriptional or post-translational variant thereof, wherein the biomarkers are present in urine. Another embodiment of the invention relates to the use of peptide sequences derived from APOAl, GSN40, GSN80 and TTHY or a transcriptional or post-translational variant thereof, wherein the biomarkers are present in urine. Another embodiment of the invention relates to the use of peptide sequences derived from AM ?, APOAl, GSN40 and IDHC or a trans-transcriptional or post-translational variant thereof, wherein the biomarkers are present in urine. Another embodiment of the invention relates to the use of peptide sequences derived from CATD H, ENOA, GSTP1 and PRDX2 or a transcriptional or post-translational variant thereof, wherein the biomarkers are present in urine. Another embodiment of the invention relates to the use of peptide sequences derived from CATD K, ENOA, PRDX2 and TTHY or a transcriptional or post-translational variant thereof, wherein the biomarkers are present in urine. Another embodiment of the invention relates to the use of peptide sequences derived from AMY, CATD K, ENOA, IDHC and PRDX2 or a transcppcional or post-translational variant thereof, where biomarkers are present in urine. Another embodiment of the invention relates to the use of peptide sequences derived from AMY, APOAl, GSN40, GSN80 and TTHY or a transcriptional or post-translational variant thereof, wherein the biomarkers are present in urine. Another embodiment of the invention relates to the use of peptide sequences derived from CATD H, ENOA, GSTP1, MASP2, PRDX2 and TTHY or a transcriptional or post-translational variant thereof, wherein the biomarkers are present in urine. Another embodiment of the invention relates to the use of peptide sequences derived from AMY, APOAl, CATD K, ENOA, IDHC, PRDX2 and TTHY or a transcriptional or post-translational variant thereof, wherein the biomarkers are present in urine. Another embodiment of the invention relates to the use of one or more peptide sequences derived from the biomarkers selected from AMY, APOAl, GSN40 and GSN80 or a transcriptional or post-translational variant thereof and one or more peptide sequences derived from selected biomarkers of TTHY, ACY1, AKR1A1, ALDOB, ANXA4, BIEA, BLVRB, CATD H, CATD K, C03, CPGL1, ENOA, FRIL, GDIB, GPX3, GSHB, GSTPl, IDHC, LMAN2, LY6G6E, MASP2, NADC, NAPSA, PA2G4, PARK7, PCBP1, PDC6I, PPAL, P, PTD012, QPCT, SBPl, STIP1, TALDO, WDRl and PETBP or a transentracional or post-translational variant thereof, in which the biomarkers are present in urine. Another embodiment of the invention relates to the use of one or more peptide sequences derived from biomarkers selected from AMY, APOAl, GSN40 and GSN80 or a trans-transcriptional or post-translational variant thereof and one or more peptide sequences derived from selected biomarkers of TTHY, ACY1, AKR1A1, ALDOB, ANXA4, BIEA, BLVRB, CATD H, CATD K, C03, CPGL1, ENOA, FRIL, GDIB, GPX3, GSHB, GSTP1, IDHC, LMAN2, LY6G6E, MASP2, NADC, NAPSA, PA2G4 , PARK7, PCBP1, PDC6I, PPAL, P, PTD012, QPCT, SBP1, STIP1, TALDO and WDR1 or a transcriptional or post-translational variant thereof, wherein the biomarkers are present in urine. Another embodiment of the invention relates to the use of one or more peptide sequences derived from the selected biomarkers of CATD H, CATD K, ENOA, GSHB, GSTP1, IDHC, PPDX2 and TTHY or a transcriptional or post-translational variant thereof and one or more peptide sequences derived from biomarkers selected from ACY1, AKR1A1, ALDOB, ANXA4, BIEA, BLVRB, C03, CPGL1, FRIL, GDIB, GPX3, LMAN2, LY6G6E, MASP2, NADC, NAPSA, PA2G4, PARK7, PCBP1, PDC6I , PPAL, PTD012, QPCT, SBP1, STIP1, TALDO, WDR1, AMY, APOAl, GSN40, GSN80 and RETBP or a transcriptional or post-translational vanant thereof, wherein the biomarkers are present in urine. Another embodiment of the invention relates to the use of one or more peptide sequences derived from selected biomarkers of THTH, CATD H, CATD K, ENOA, GSTP1, IDHC, MASP2, P, AMY, APOAl, GSN40, GSN80 and RETBP or a transcriptional or post-translational variant thereof, wherein the biomarkers are present in urine. Another embodiment refers to a device for carrying out a method as defined previously comprising 1) any combination of antibodies that specifically recognizes one or more of these proteins and 2) a vehicle in a suitable package, the equipment being used to detect the presence of BTCC, to determine the stage or severity of this cancer, to evaluate the lack of disease after surgical resection, to establish the diagnosis and / or prognosis of this cancer and / or to monitor the effect of the treatment administered to an individual who suffers from said cancer. The kit may comprise a container for housing at least one agent that binds a biomarker present in a urine sample; and instructions for using at least one agent to determine the status of BTCC. Such equipment may comprise a vehicle, package or container having compartments for receiving one or more containers such as vial, tubes, and the like, each of the containers comprising one of the separate elements that will be used in the method. For example, the containers can comprise a sound that is or can be marked and also be detected. The probe can be an antibody or polynucleotide specific for a bladder cancer protein or a bladder cancer gene, respectively. Alternatively, the equipment may comprise a mass spectrometry (MS) probe. The kit may also include containers containing nucleotides for amplification or silencing of a target nucleic acid sequence, and / or a container comprising a reporter medium, such as a biotin binding protein, e.g., avidite or streptavidin, linked to a detectable label, e.g., an enzymatic, fluorescent or radioisotopic label. The kit may include another or part of the amino acid sequence of biomarkers of bladder cancer or a nucleic acid molecule encoding said amino acid sequences. The equipment of the invention will normally comprise the container described above and one or more containers comprising materials suitable from a commercial and user's point of view., including regulatory solutions, diluents, filters, needles, syringes and packaging inserts with instructions for use. In addition, a label may be provided in the container to indicate the composition that is used for a specific therapeutic or non-therapeutic application and may also include instructions for use, such as those described above. Instructions and / or other information may also be included in an insert that is included with the equipment. Another embodiment refers to a device for carrying out a method as previously defined, comprising a biomicrocircuit. Another embodiment refers to a device for carrying out a method as previously defined comprising a biomicrocircuit, wherein the biomicrocircuit comprises antibodies for the detection of one or more biomarkers selected from 40 biomarkers of bladder cancer or a trans-translational or post-translational variant of the biomarker. same. There is a wide range of immunological assays available to detect and / or quantify the formation of competitive or non-competitive antigen-antibody complexes.; previously, numerous competitive or non-competitive protein binding assays have been described and a large number thereof are commercially available. Therefore, the proteins can be quantified with antibodies such as, for example. Monoclonal antibodies, polyclonal antibodies, either intact or recombinant fragments thereof, body-combis and Fab or scFv fragments of antibodies, specific for proteins. These antibodies can be of human origin, humanized or of animal origin. On the other hand, they can be marked or unmarked and can be used in a wide range of analysis. Marker molecules that can be used to label antibodies include radionuclides, enzymes, fluorophores, chemiluminescent reagents, enzyme substrates or co-factors, enzyme inhibitors. Particles, colorants and derivatives. While the specificity of antibody binding is greater, the antigen concentration that can be detected is lower. There is a wide variety of assays well known to those skilled in the art that can be used in the present invention, which uses unlabeled antibodies (primarily antibodies) and labeled antibodies (secondary antibodies); These techniques include but are not limited to Western analysis or Western blotting, ELISA (Enzyme Linked Immunosorbent Assay), RIA (Radiommunoanalysis), competitive EIA (competitive enzyme immunoassay), DAS-ELISA (double-antibody sandwich ELISA), techniques immunocytochemistry or immunohistochemistry, techniques based on the use of biomicrocircuits of microdissections of proteins that include specific antibodies, analysis based on colloidal gold precipitation in formats such as submersible strips; or by affinity chromatography techniques, ligand binding analysis or lectin binding analysis. Preferred embodiments of this aspect of the invention are protein micro-arrangements and double-antibody sandwich ELISA (DAS-ELISA). In this immunoassay, any antibody, or combination of antibodies, which are specific against one or more epitopes of the proteins of the invention can be used. As an example of one of the many possible formats of this analysis, a monoclonal or polyclonal antibody or a fragment of this antibody or a combination of those antibodies that recognizes one or more epitopes of the 40 proteins of the invention are connected to the surface of a solid phase support and placed in contact with the sample that will be analyzed and incubated for a specific time and under appropriate conditions to form the antigen-antibody complexes. After washing under appropriate conditions to remove nonspecific complexes, a reporter reagent, consisting of a monoclonal or polyclonal antibody, or a fragment of this antibody, or a combination thereof and recognizing one or more epitopes of the the invention, bound to a molecule that generates a signal, is incubated with the antigen-antibody complexes under appropriate conditions of time and temperature. The presence of one or more proteins selected from the proteins of the invention in the sample to be analyzed was detected and quantified and the signal generated was measured. In order to avoid signal variation due to differences in total protein concentration between the samples, all measurements are normalized. As indicated above, the method of the invention involves monitoring the stage of bladder carcinoma by quantifying the differentially expressed soluble proteins within a urine sample through specific antibodies. As will be appreciated by those skilled in the art, any means for specifically identifying and quantifying these proteins are contemplated. In the following description, the generalized method for obtaining and analyzing the total protein content of human urine samples (here referred to as the sample) is described. The method involves processing the samples, the use of 2D electrophoresis to separate proteins within the sample, the selection of differentially expressed proteins by means of image and static analysis of different samples and the use of one or more of the differentially expressed proteins of the invention to generate specific antibodies for proteins that will be used as markers for bladder cancer. Comparative protein analyzes were carried out between samples obtained from healthy individuals (controls) and patients diagnosed with BTCC (Ta, low IT grade, high IT grade, and 12). In an attempt to identify proteins differentially expressed in different stages of bladder cancer and during the progression of bladder cancer. Among all the proteins that showed differential expression, 40 proteins changed more than twice their reproductive capacity. They were identified by peptide mass fingerprint using mass spectrometry and database search. The present invention will be further illustrated by the following examples. These examples are given by way of illustration only and are not considered to be limiting.

I. Identification of differentially expressed proteins To identify differentially expressed proteins throughout the progression of bladder cancer, protein profiles of healthy urine were compared with those of patients with early and advanced stage bladder tumor using a proteomic approach. Urine samples (150 in total) were collected from healthy individuals and patients with transitional bladder cancer by visiting the urology units of the Hospitals that belong to the Spanish Public Health Network. These samples were classified as follows: a) Without carcinoma (63 samples) including: Controls (CV, 32 samples!: Patients who had bladder cancer and had tumor resection and were controlled (cystoscopy showed no other affectations). - Patients with other diseases of the urological tract (31 samples, including benign prostatic hyperplasia, prostate cancer) b) BTCC (87 samples): patients diagnosed with the disease at different stages of development including: - Ta (21 samples) ) - Low-grade IT (29 samples) - High-grade IT (19 samples) - T2 (18 samples) The BTCC group in the sample was accompanied by a biopsy that was the key to its classification in the stages of cancer development of bladder A Process of urine samples and protein separation The urine samples were frozen at -80 ° C and shipped to the laboratory on dry ice without rupture of the cold chain. The samples were kept at -80 ° C until they were processed. The samples were very heterogeneous, varying from light yellow to red urine with blood clots, transparent or containing suspended tissues. The total volume was also very variable from 5 to 100 ml. The protein concentration of the samples ranged from 20 μg / ml to 2 mg / ml (150 μg / ml was the average concentration), the protein content was obtained, samples were thawed on ice and centrifuged at 2000 μg for 5 min at 4 ° C. The supernatant was used to determine the protein concentration The volume needed to precipitate 100 μg of protein was calculated taking into account that the precipitation efficiency of protein with 15% w / v trichloroacetic acid (TCA ) is 75% The rest of the urine sample was frozen again at -80 ° C and stored for additional 2D electrophoresis (if necessary) TCA and urine were mixed for 1 hour on ice and then centrifuged at 16000 μg for 20 in at 4 ° C to obtain pellet of precipitated proteins This pellet was washed with acetone, stored at -20 ° C and dried by evaporation of solvent To carry out two-dimensional electrophoresis experiments (2D ), the first dimension was IEF (e isoelectric focus), where proteins are separated by their charge (pl); the second dimension was SDS-PAGE where the proteins were separated by their molecular weight. To carry out the first dimension, the dried protein pellet was resuspended with 450 μl of rehydration buffer (Urea 7M, 2M thiourea, 2% CHAPS, 2% IPG buffer, bromophenol blue 0.002%) for 1 hour at room temperature. The IPG buffer (Amersham, ref. # 17-600-88) was used so that IEF varied from pH 3-10. For IEF, the Etmers ™ IPGphor ™ Isoelectric Focusing System from Amersham was used according to the manufacturer's instructions. IEF was carried out on immobilized pH gradients, named IPG strips, purchased from Amersham (Ref # 17-6002-45). The solubilized proteins focused in the first dimension on the strips after 16 hours of active rehydration of the gel at 30B. Then the voltage was increased to 8000 V, the intensity was never higher than 50 μA per gel. IEF ended when the voltage reached 90000 Vhz. For the second dimension, 26 x 20 cm 12.5% acrylamide was polymerized in the laboratory using Castor Gel twelve Ettan DALT from Amersham. The gels were operated on the Large Format Vetical System twelve DALT and following the manufacturer's instructions to the left front end of gel electrophoresis. The eles are colored silver nitrate using the Amersham coloring equipment (17-1150-01) following the manufacturer's instructions. The gels were dried and stored for analysis of subsequent images of the protein spots (Figure IA). For some urine samples, more than one 2D gel was operated.

B. Analysis of the protein points in which each gel was scanned to obtain the map of points for image analysis. Progenesis PG220 software from Nonlinear Dynamics (UK) was used to analyze image files in a format of 300 dpi (dot per inch) and 8 bit / channel. To increase the resolution, the analyzes were carried out in discrete areas of gels. For each area, named as A (GIGURA 2B), K (FIGURE 1C), R (FIGURE ID) and S (FIGURE 1E), the best gels were selected and scanned for image analysis. Progenesis PG220 software transformed the information of the flat image into a 3D image, where the intensity of each point was correlated with its volume and with the relative amount of the corresponding protein in the individual's urine. With the software tables, the intensities of each point were obtained in each gel. These raw data were the basis for the subsequent statistical analyzes. To perform the statistical analyzes of each individual point and compare its intensity in two different groups (for example, CV and Ta), a normal distribution had to be tested in these data blocks. To compare the points belonging to two groups, a Student t test was applied and a p-value was obtained: this was the theoretical error to assign a point to one subgroup or another. Only points with values p = 0.05 were selected. The average field or relation of these points was calculated, each one being called 1) the total volume of points of the sample, that is, the theoretical total protein content, or 2) the protein expressed constantly, the amount of which is constant in each sample,? 3) a second protein differentially expressed from the same sample. Also the value n or number of samples was taken into account. 40 proteins showed statistical significance and strength in their change of times when compared with urine samples from control patients with urine samples from patients diagnosed with transitional bladder cancer. The identification of the points was carried out by MALDI-TOF spectroscopy (Time of Flight of Desorption / Ionization of Laser Assisted by Matrix). The urine samples whose 2D electrophoresis showed statistically significant spots underwent re-electrophoresis 2D and the points taken from the gel. The proteins were digested and analyzed by the MALDI-TOF spectrometer. The fingerprint of the peptide mass allowed the identification of 40 proteins, which were identified as TTHY, ACY1, AKR1A1, ALDOB, ANXA, BIEA, BLVRB, CATD H, CATD K, C03, CPGL1, ENOA, FRIL, GDIB, GPX3 , GSHB, GSTPl, IDHC, LMAN2, LY6G6E, MASP2, NADC, NAPSA, PA2G4, PARK7, PCBP1, PDC6I, PPAL, PRDX2, PTD012, QPCT, SBPl, STIP1, TALDO, WDRl, AMY, APOAl, GSN40, GSN80 and RETBP (see table 1). Therefore, these proteins, which were shown to be differentially expressed in patients diagnosed with BTCC, were identified as biological markers of significant value for the diagnosis, prognosis, monitoring and treatment of the disease.

Table 1 As an example, Figure 2 shows the R211 point corresponding to BIEA in the R area of the two-dimensional electrophoresis (2D) gel obtained from urine samples from a healthy individual (Figure 2A), of a patient suffering from cancer in the Ta stage (Figure 2B), of a patient suffering from cancer at the stage of low IT grade (Figure 2C), of a patient suffering from cancer at the stage of high TI grade (Figure 2D) and from a patient suffering from cancer in stage T2 (Figure 2E). It can be observed that the intensity of the R211 point is clearly decreased in the samples of cancer patients in different stages, when compared with the healthy urine sample. It was shown that these important differences after statistical analysis with Progenesis PG220 software. Figure 3 shows the intensity of point R211 in the CV samples. Ta, low Ti grade, high IT grade and T2. The number of samples for each group is indicated in parentheses. Figure 4 shows the strength of intensity measurement of point R211 in different urine samples. This is expressed as the percentage of the gels that maintain the presence or absence of the R211 point in each gel analyzed. The number of samples for each group again is indicated in parentheses. Table 2 shows p and the values of changing times of the statistical analysis for point R211 in the samples from different stages of cancer compared to a sample from an individual without cancer (CV). The change of times of the R211 point was normalized by the total point volume (TSV) for each individual gel.

II. Development of antibody for the proteins found in the urine sample The available polyclonal and / or monoclonal antibodies (either commercially purchased or generated following the immunization protocol) were tested for reactivity and sensitivity, the method generally involving the preparation of a standard curve ("dose response") for the protein that will be monitored.

A. Immunization Protocols 1. Polyclonal Antibodies Polyclonal antisera that may arise against a given protein using standard methodologies, such as those described in numerous texts available in the art and known to those skilled in the art. In this example, New Zealand white male rabbits were immunized with the protein preparations first in Freund's complete adjuvant (Gibco, Grand Island, N.Y.) and each month with the protein with incomplete adjuvant for three months. The rabbit sera and serum from mice before the fusion are used as polyclonal antiserum and are shown by the normal Western analysis technique to be reactive with the protein preparations. 2. Generation of Recombinant Proteins The proteins found in the gels in very low amounts for immunization were cloned and expressed in E. coli using the cloning vector? ET28b (+). Crude extracts were obtained as previously described (Boronat A., et al., J. Bacterial, 1981, 147: 181-85) and run on an SDS-PAGE gel. For the generation of antibodies against recombinant proteins, the released proteins are used directly to immunize rabbits as described above.

B. Western Analysis Experiments Protein samples (20 μg total protein) were mixed with SDS-PAGE gel loading buffer solution with 5% β-mercaptoethanol and incubated at 100 ° C for 5 minutes, before being loaded in 6-f polyacrylamide gel. The electrophoresis proteins were then transferred to nitrocellulose membranes. The gels in duplicate were run and plotted. A membrane was tested with antibodies raised against one or more of the selected proteins of the invention (Santa Cruz Biotech, Inc., Santa Cruz, CA, USA) while testing the second membrane with an antibody raised against actin (Amersham, Little Chalfont, UK) as a control to load protein. Finally, the membranes were hybridized with a secondary antibody conjugated with peroxidase (Amersham) and the chemiluminescent signal was detected using the ECL system (Amersham) with high performance chemilummization film (Hyperfilm ECL, Amersham).

III. Validation of urine test samples using Western analysis experiments Several biomarkers were tested on 340 blind urine samples comprising: a) Without Carcinoma (12 samples) b) BTCC including: Ta (8 samples), low IT grade (6 samples) ), high grade IT (6 samples) and T2 (8 samples) The values obtained for sensitivity, specificity and precision are shown in Table 3. The sensitivity, specificity and precision values were calculated using these formulas: (real negatives + real positives) Accuracy = Number of samples (real positives) Accuracy = Number of samples (real negatives) Accuracy = Number of samples For example, the AMY biomarker shows sensitivity of 89% and specificity of 75o for a precision value of 85 °.

Table 3 By a binomial logistic regression of the data obtained in these experiments it was possible to obtain different combinations of biomarkers that generally increased the sensitivity and specificity of the diagnostic method, as shown in table 4.

Table DESCRIPTION OF THE DRAWINGS Figures IA to 1E: Two-dimensional electrophoresis gel (2D) obtained from a urine sample and showing the four different areas studied A, K, R and S (Figure IA), area A (Figure IB), area K (Figure 1C), AREA r (Figure ID) and area S (Figure 1E) Figures 2A to 2E: Point R211 corresponding to BIEA in the R area of two-dimensional (2D) electrophoresis gel obtained from urine samples of a healthy individual (Figure 2A), of a patient suffering from cancer in the stage Ta (Figure 2B), of a patient suffering from cancer in the low-grade stage of IT (Tl-LG, Figure 2C) of a patient suffering from cancer in the high-grade stage of IT (Tl-HG, Figure 2D) and a patient suffering from cancer in stage T2 (Figure 2E). Figure 3: Point intensity R211 in the samples of CV, Ta, low degree of IT (Tl-LG), high degree of TI (Tl-HG) and T2. The number of samples for each group is indicated in parentheses. Figure 4: The strength of the intensity measurement for the R211 point in different urine samples. This is expressed as the percentage of the gels that maintain the presence or absence of the R211 point in each gel analyzed. The number of samples for each group is indicated in parentheses.

Claims (44)

1. - A non-invasive method consists of: a) detecting and quantifying at least two biomarkers in a sample of an individual's urine test; where at least two of the biomarkers are selected from a group consisting of TTHY (SEQ ID 1),

ACY1 (SEQ ID 2), AKR1A1 (SEQ ID 3), ALDOB (SEQ ID 4), ANXA4

(SEQ ID 5), BIEA (SEQ ID 6), BLVRB (SEQ ID 7), CATD H (SEQ ID 8), CATD K (SEQ ID 9), C03 (SEQ ID 10, SEQ ID 11), CPGL1 (SEQ. ID 12), ENOA (SEQ ID 13, SEQ ID 14), FRIL (SEQ ID 15), GDIB (SEQ ID 16), GPX3 (SEQ ID 17), GSHB (SEQ ID 18), GSTPl (SEQ ID 19), IDHC (SEQ ID 20), LMAN2 (SEQ ID 21), LY6G6E (SEQ ID 22), MASP2 (SEQ ID 23), NADC (SEQ ID 24), NAPSA (SEQ ID 25), PA2G4 (SEQ ID 26), PARK7 (SEQ ID 27), PCBP1 (SEQ ID 28), PDC6I (SEQ ID 29), PPAL (SEQ ID 30), PRDX2 (SEQ ID 31), PTD012 (SEQ ID 32), QPCT (SEQ ID 33), SBPl ( SEQ ID 34), STIP1 (SEQ ID 35), TALDO (SEQ ID 36), WDRl (SEQ ID 37), AMY (SEQ ID 38, SEQ ID 39, SEQ ID 40), APOAl (SEQ ID 41), GSN40 ( SEQ ID 42), GSN80 (SEQ ID 43) and RETBP (SEQ ID 44) or a transcriptional or transnational variant thereof; and b) comprises the measure of expression obtained in: a) In the urine test sample at the corresponding standard value in the urine test sample, where variations in the measure obtained in a) compared to the corresponding standard value in the Normal urine is indicative of carcinoma of the transition cell of the vejiga. 2. The method according to claim 1, wherein step a) comprises detecting and quantifying at least two biomarkers selected from TTHY, ACY1, AKR1A1, ALDOB, ANXA4, BIEA, BLVRB, CATD H, CATD K, C03 , CPGL1, ENOA, FRIL, GDIB, GPX3, GSHB, GSTPl, IDHC, LMAN2, LY6G6E, MASP2, NADC, NAPSA, PA2G4, PARK7, PCBP1, PDC6I, PPAL, PRDX2, PTD012, QPCT, SBPl, STIP1, TALDO and WDRl or a transcppcional or post-translational variant thereof. 3. The method according to claim 1, wherein step a) comprises detecting and quantifying at least two biomarkers selected from TTHY, AMY, APOAl, GSN40 and GSN80 or a transcriptional or post-translational variant thereof.

4. The method according to claim 1, wherein step a) comprises detecting and quantifying at least two biomarkers selected from AMY, APOAl, GSN40 and GSN80 or a transcriptional or post-translational variant thereof and one or more selected biomarkers of TTHY, ACY1, AKR1A1, ALDOB, ANXA4, BIEA, BLVRB, CATD H, CATD K, C03, CPGL1, ENOA, FRIL, GDIB, GPX3, GSHB, GSTP1, IDHC, LMAN2, LY6G6E, MASP2, NADC, NAPSA , PA2G4, PARK7, PCBP1, PDC6I, PPAL, PRDX2, PTD012, QPCT, SBP1, STIP1, TALDO and WDR1 or a transcppcional or post-translational variant thereof.

5. The method according to claim 1, wherein step a) comprises detecting and quantifying at least two biomarkers selected from CATD H, CATD K, ENOA, GSHB, GSTP1, IDHC, PRDX2 and TTHY or a transcriptional variant or post-translational thereof and one or more selected biomarkers of ACY1, AKR1A1, ALDOB, ANXA4, BIEA, BLVRB, C03, CPGL1, FRIL, GDIB, GPX3, LMAN2, LY6G6E, MASP2, NADC, NAPSA, PA2G4, PARK7, PCBP1 , PDC6I, PPAL, PTD012, QPCT, SBP1, STIP1, TALDO, WDR1, AMY, APOAl, GSN40, GSN80 and RETBP or a transcriptional or post-translational variant thereof.

6. The method according to claim 1, wherein step a) comprises detecting and quantifying at least two biomarkers selected from TTHY, CATD H, CATD K, ENOA, GSTP1, IDHC, MASP2, PRDX2, AMY, APOAl , GSN40, GSN80 and RETBP or a transcriptional or post-translational variant thereof.

7. - The method according to claims 1 to 6, wherein step a) comprises detecting and quantifying at least three biomarkers selected from the group as defined in claim 1.

8. The method according to the claims 1 to 6, wherein step a) comprises detecting and quantifying at least four biomarkers selected from the group as defined in claim 1.

9. The method according to claims 1 to 6., wherein step a) comprises detecting and quantifying at least five biomarkers selected from the group as defined in claim 1.

10. The method according to claim 1, wherein step a) comprises detecting and quantifying the biomarkers APOAl, GSN40 and TTHY or a transcriptional or post-translational variant thereof.

11. The method according to claim 1, wherein step a) comprises detecting and quantifying the biomarkers APOAl, GSN40, GSN80 and TTHY or a transcriptional or post-translational variant thereof.

12. The method according to claim 1, wherein step a) comprises detecting and quantifying the biomarkers APOAl, GSN40, GSN80 and TTHY or a transcriptional or post-translational variant thereof.

13. The method according to claim 1, wherein step a) comprises detecting and quantifying the biomarkers CATDH, ENOA, GSTP1, MASP2, PRDX2 and TTHY or a transcriptional or post-translational variant thereof.

14. The method according to claim 1, wherein step a) comprises detecting and quantifying the biomarkers AMY, APOAI, CATD K, ENOA, IDHC, PRDX2 and TTHY or a transcriptional or post-translational variant thereof.

15. The method according to any of claims 1 to 14, are used to detect the presence of carcinoma of the transition cell of the bladder, to determine the season or severity of this cancer, to assess the lack of the disease after surgical resection, to establish the diagnosis and / or prognosis of this cancer and / or to monitor the effect of the treatment administered to the individual suffering from said cancer.

16. The method according to claims 1 to 15, wherein the urine sample to be analyzed was obtained from an individual previously undiagnosed with carcinoma of the transitional cells of the bladder.

17. The method according to claims 1 to 15, wherein the urine sample to be analyzed was obtained from an individual previously diagnosed with carcinoma of the transitional cells of the bladder.

18. The method according to the claims 1 to 15, wherein the urine sample to be analyzed was obtained from an individual receiving a treatment against carcinoma of the transitional cells of the bladder.

19. The method of any of claims 1 to 15, characterized in that the detection and quantification of the biomarkers comprises the first step, which the protein extract of the urine sample is contacted with the composition of at least two antibodies specific to one or more epitopes of the biomarkers of claim 1, and the second step, in which the resulting protein-antibody complexes are quantified.

20. The method according to claim 19, characterized in that said antibodies are of human origin, humanized or of non-human origin and selected from monoclonal or polyclonal antibodies, intact or recombined fragments of antibodies, combined bodies and fragments of Fab antibodies. or scFv.

21. The method according to claim 19 or 20, characterized in that in the detection and quantification of the resulting protein-antibody complexes, the techniques used are selected from a group consisting of: Western analysis, ELISA (Immunoabsorbant Assay of linked enzymes), Competitive EIA (Competitive Enzyme Immunoassay), DAS-ELISA (double-antibody sandwich ELISA), immunocytochemical or immunohistochemical techniques, techniques based on the use of biomicrocircuits or protein micro-rays that include specific antibodies, assays based on precipitation of colloidal gold in formats such as submersible strips; or by affinity chromatographic techniques, ligand binding assays or lectin binding assays.

22. The method according to any of claims 1 to 21, wherein the method is used to monitor the efficacy of pharmacological or surgical treatments administered to an individual suffering from transitional cell carcinoma of the bladder.

23. The method according to claims 1 to 22, wherein the method is used to determine the progression of the disease when at least two biomarkers are compared from different urine samples from the same patient obtained at different times within the evolution of bladder transition cell carcinoma.

24. The use In Vi tro of at least two sequences of peptides derived from the biomarkers as defined in claim 1, in the urine to detect the presence of transition cell carcinoma of the bladder by urinalysis, to determine the season or severity of this cancer, to assess the lack of the disease after surgical resection, to establish the diagnosis and / or prognosis of this cancer and / or to monitor the effect of the treatment administered to the individual suffering from said cancer .

25. The use according to claim 24 of at least two peptide sequences derived from the biomarkers selected from TTHY (SEQ ID 1), ACY1 (SEQ ID 2), AKR1A1 (SEQ ID 3), ALDOB (SEQ ID NO. 4), ANXA4 (SEQ ID 5), BIEA (SEQ ID 6), BLVRB (SEQ ID 7), CATD H (SEQ ID 8), CATD K (SEQ ID 9), C03 (SEQ ID 10, SEQ ID 11) , CPGL1 (SEQ ID 12), ENOA (SEQ ID 13, SEQ ID 14), FRIL (SEQ ID 15), GDIB (SEQ ID 16), GPX3 (SEQ ID 17), GSHB (SEQ ID 18), GSTPl (SEQ. ID 19), IDHC (SEQ ID 20), LMAN2 (SEQ ID 21), LY6G6E (SEQ ID 22), MASP2 (SEQ ID 23), NADC (SEQ ID 24), NAPSA (SEQ ID 25), PA2G4 (SEQ ID NO. 26), PARK7 (SEQ ID 27), PCBP1 (SEQ ID 28), PDC6I (SEQ ID 29), PPAL (SEQ ID 30), PRDX2 (SEQ ID 31), PTD012 (SEQ ID 32), QPCT (SEQ ID 33) ), SBPl (SEQ ID 34), STIP1 (SEQ ID 35), TALDO (SEQ ID 36), WDRl (SEQ ID 37), AMY (SEQ ID 38, SEQ ID 39, SEQ ID 40), APOAl (SEQ ID 41) ), GSN40 (SEQ ID 42), GSN80 (SEQ ID 43) and RETBP (SEQ ID 44) or a transcriptional or post-translational variant thereof.

26. The use according to claim 25, of at least two peptide sequences derived from the selected biomarkers of TTHY, ACY1, AKR1A1, ALDOB, ANXA4, BIEA, BLVRB, CATD H, CATD K, C03, CPGL1, ENOA, FRIL, GDIB, GPX3, GSHB, GSTP1, IDHC, LMAN2, LY6G6E, MASP2, NADC, NAPSA, PA2G4, PARK7 , PCBP1, PDC6I, PPAL, PRDX2, PTD012, QPCT, SBP1, STIP1, TALDO and WDR1 or a transcriptional or post-translational variant thereof.

27. The use according to claim 25, of at least two peptide sequences derived from the biomarkers selected from TTHY, AMY, APOAl, GSN40 and GSN80 or a transcriptional or post-translational variant thereof.

28. The use according to claim 25, of one or more polypeptide sequences derived from biomarkers selected from AMY, APOAl, GSN40 and GSN80 or a trans-transcriptional or post-translational variant thereof and one or more derived peptide sequences of selected biomarkers of TTHY, ACY1, AKR1A1, ALDOB, ANXA4, BIEA, BLVRB, CATD H, CATD K, C03, CPGL1, ENOA, FRIL, GDIB, GPX3, GSHB, GSTP1, IDHC, LMAN2, LY6G6E, MASP2, NADC , NAPSA, PA2G4, PARK7, PCBP1, PDC6I, PPAL, PRDX2, PTD012, QPCT, SBP1, STIP1, TALDO and WDR1 or a transcppcional or post-translational variant thereof.

29. The use according to claim 25, of one or more peptide sequences derived from the selected biomarkers of CATD H, CATD K, ENOA, GSHB, GSTP1, IDHC, PRDX2 and TTHY or a transcriptional or post-translational variant thereof and one or more peptide sequences derived from the selected biomarkers of ACY1, AKR1A1, ALDOB, ANXA4, BIEA, BLVRB, C03, CPGL1, FRIL, GDIB, GPX3, LMAN2, LY6G6E, MASP2, NADC, NAPSA, PA2G4, PARK7 , PCBP1, PDC6I, PPAL, PTD012, QPCT, SBP1, STIP1, TALDO, WDR1, AMY, APOAl, GSN40, GSN80 and RETBP or a transcriptional or post-translational variant thereof.

30. The use according to claim 25, of at least two peptide sequences derived from the selected biomarkers of TTHY, CATD H, CATD K, ENOA, GSTP1, IDHC, MASP2, PRDX2, AMY, APOAl, GSN40, GSN80 and RETBP or a transcriptional or post-translational variant thereof.

31. The use according to the claim of any of claims 25 to 30, wherein the variations in the measurements of at least two biomarkers in the sample of the urine test in comparison with the corresponding standard values in Normal urine is indicative of the transition cell carcinoma of the bladder.

32. The use according to any of claims 25 to 30, of at least three peptide sequences.

33. The use according to claims 25 to 30, of at least four peptide sequences.

34. The use according to claims 25 to 30, of at least five peptide sequences. 35.- The use according to claim 25, of the sequences of peptides derived from APOAl, GSN40 and TTHY or a transcriptional or post-translational variant thereof. 36. The use according to claim 25, of the sequences of peptides derived from APOAl, GSN40, GSN80 and TTHY or a transcriptional or post-translational variant thereof. 37. The use according to claim 25, of the peptide sequences derived from AMY, APOAl, GSN40, GSN80 and TTHY or a transcriptional or post-translational variant thereof. 38.- The use according to claim 25, of the peptide sequences derived from CATD H, ENOA, GSTP1, MASP2, PRDX2 and TTHY or a transcriptional or post-translational variant thereof. 39. The use according to claim 25, of the peptide sequences derived from CATD H, ENOA, GSTP1, MASP2, PRDX2 and TTHY or a transcriptional or post-translational variant thereof. 40.- The use of one or more nucleotides or sequences of peptides derived from one or more biomarkers selected from the group as defined in claim 1, alone or in any combination, in methods for displaying, identifying, developing and / or evaluating the efficacy of therapeutic agents to treat bladder transition cell carcinoma. 41. A package for carrying out a method as defined in any of claims 1 to 23, comprising 1) at least two antibodies that specifically recognize independently one of the biomarkers of claim 1 and 2) a race in an adequate package. 42. A package according to claim 41, which is used to detect the presence of transitional cell carcinoma of the bladder, to determine the season or severity of this cancer, to assess the lack of the disease after surgical resection , to establish the diagnosis and / or prognosis of this cancer and / or to monitor the effect of the treatment administered to the individual suffering from said cancer. 43.- A package according to claims 41 or 42, comprising a biomicrocircuit. 44. A package according to claim 43, wherein the biomicrocurcuit comprises antibodies for the detection of biomarkers selected from a group as defined in claim 1.